الفهرس 
Phytoglobins in plants
Phytoglobins and redox status in plantsOnly 14 pages are availabe for public view
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Abstract
Phytoglobins (Pgbs) are a group of plant hemoglobins that are effective scavengers of nitric oxide (NO), therefore, they interfere with many NOmediated plant responses to stress, including hypoxia. Low oxygen levels, experienced by hypoxic roots, retard growth and can lead to the deterioration of the root apical meristem (RAM) by altering the functionality of the quiescent center (QC). The main function of the QC, which is located at the center of the RAM and comprised of slowly diving cells, is to act as an “organizing center” preventing the differentiation of the adjacent initials through rapid divisions. Functionality and state of quiescence in the QC is conferred by the PIN-mediated basipetal flow of auxin, which generate a highly-oxidized ascorbate environment. In this study, three maize lines, the wild type (WT), phytoglobinoverexpressed line }ZmPgb1.1 (S){ , and phytoglobin-downregulated line }ZmPgb1.1 (A){ were used. To evaluate the role of phytoglobin (ZmPgb1.1 gene expression) in preserving the root growth during hypoxia stress, these three lines were subjected to hypoxic treatment (4% oxygen) for three time points (0 h, 6 h & 12 h) and many experiments were done. Firstly, using quantitative RT-PCR, the expression of auxin biosynthetic, transporters and responsive genes in three root segments (starting from the root tip upwards, S1= 0-2 mm, S2 = 2-5 mm, and S3 = 5-10 mm.) of three maize lines were measured. The results showed that the seven auxin biosynthetic genes (ZmAO1, ZmVT2, ZmTSB1, ZmTSB2, ZmNIT1, ZmNIT2, and ZmAMI1), and one auxin responsive gene (ZmGH3) were upregulated in maize root segments of ZmPgb1.1(S) (phytoglobin overexpressed) line after 6 h of hypoxic treatment particularly in the root tip tissues (S1= 0-2 mm) compared Summary 137 to WT and ZmPgb1.1 (A) (phytoglobin downregulated) line. In addition, the expression of the auxin transporter genes (ZmPIN1 and ZmAUX1) was upregulated at the root tip of ZmPgb1.1(S) line after 12 h of hypoxic treatment. Moreover, relative expression of six genes of Auxin Responsive Factors (ZmARFs) family, one key transcription factor linked to auxin production, PLETHORA1 (ZmPLT1) and ZmDR5 responsive gene in root compartments (QC, PM & RC) of the maize lines were determined by qRT-PCR over the 0h, 6h, and 12h of the hypoxic treatment. The results showed that all these genes were upregulated in maize root of ZmPgb1.1(S) during hypoxia. The previous results were confirmed with the localization patterns of ZmPIN1a by RNA in situ hybridization technique along the RAM and Immunolocalization of IAA in (QC, PM and RC). Taken together, these results confirm that overexpression of ZmPgb1.1 maintains auxin (IAA) flow and transport during hypoxia stress in the maize root tip and consequently sustains root growth. These phenotypes is most likely due to the role of Pgbs to scavenge NO during hypoxia stress, so to verify that the phenotypes observed by altering ZmPgbs are mediated by NO, where NO content was manipulated pharmacologically in two reporter lines (ZmDR5-RFP and ZmPIN1-YFP) using the nitric oxide donors, sodium nitroprusside (SNP) and the nitric oxide scavenger, 2-4-carboxyphenyl-4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). The results showed that increased NO levels, resulting from either pharmacological treatments or by hypoxia stress, decreases auxin transport and response in the QC and PM of maize RAM of ZmPgb1.1 (A) line, resulting in root growth inhibition. Summary 138 Assay measurements of oxidized (DHA) and reduced (AA) ascorbate in the three root compartments and histochemical localization of ascorbate using AgNO3 indicated that perturbations in the expression of genes involved in auxin synthesis and transport, were more evident in hypoxic root tips down-regulating Pgbs and caused an abnormal auxin localization pattern at the root apical meristem, which alters the oxidized environment needed to preserve the integrity of the cells in the quiescent center (QC). A switch from an oxidized (DHA) to a reduced (AA) environment, with a prevalent accumulation of reduced ascorbic acid occurs in QCs suppressing Pgbs resulting in the premature differentiation of the initials and root growth retardation. Over-expression of Pgb alleviates the root growth inhibition during hypoxia by reducing NO levels and maintaining QC viability through the retention of auxin flow and the oxidized environment at the root tip. The results obtained from the relative expression measurements of ten of genes encoding ascorbate associated enzymes (ZmAAO, ZmAPX1, ZmAPX2, ZmAPX4, ZmMDHAR1, ZmMDHAR4, ZmDHAR1, ZmDHAR2 & ZmDHAR3) using quantitative RT-PCR confirmed the results of ascorbate assay and localization. Root regeneration of hypoxic QC, PM and RC cells further showed that, relative to wild type (WT), root regeneration percentage in maize plants suppressing the Pgbs }ZmPgb1.1 (A){ was highly lower than in those in which the Pgbs are over-expressed}ZmPgb1.1 (S){. In conclusion, the presence of Pgbs is needed to scavenge NO and to preserve cell viability and root integrity during conditions of hypoxia. In addition, overexpression of Pgbs is participating in root apical meristem maintenance under hypoxic stress.